Summary: Heart failure (HF) is a devastating disease with poor prognosis. Pathologic cardiac remodeling is characterized by progressive hypertrophy, apoptosis and fibrosis. Fibrosis exacerbates cardiac remodeling, arrhythmias and decreased ventricular compliance; emerging evidence suggests that cardiac fibroblasts (CFs) play a key role in these processes. To date, there are no therapies that target fibrosis. Elucidating the role(s) of CF in pathologic remodeling may hold substantial therapeutic potential. CF studies have been limited by the lack of genetic reagents in vivo. We are collaboratively characterizing two novel mouse models (PeriostinMerCreMer and Tcf21MerCreMer knock-in mice) that permit targeted, inducible manipulation of CF function/ viability to directly determine the mechanistic basis of cardiac fibrosis. Progressive cardiac fibrosis occurs in large part due to an imbalance in the production and degradation of the extracellular matrix (ECM). CFs are the major matrix-producing cells in the heart. CFs maintain ECM homeostasis; this process is significantly dysregulated in cardiac fibrosis. Transition of CFs to myofibroblasts (MF) results in excess deposition of ECM components, thus exacerbating HF. The ECM protein fibronectin (FN) plays a key role in pathologic remodeling of the ECM in HF; CFs are the major source of cellular FN in the heart. Production and polymerization of FN are elevated in clinical and experimental HF; these processes are essential for MF transition and pathologic fibrosis. FN polymerization tightly regulates the assembly of ECM proteins, including collagens type I and III; it also promotes cell adhesion, growth, migration and contractility. We recently described a peptide, pUR4, which binds to FN and inhibits its cell-mediated assembly and polymerization. pUR4 was injected IP daily for 7 days following cardiac ischemia/reperfusion (I/R) injury, followed by three more weeks of reperfusion. Our preliminary data suggest that 7 days of pUR4 post-I/R reduces FN deposition, collagen accumulation and myocardial fibrosis and significantly improves cardiac function. To test the effect of FN ablation in fibroblasts following cardiac injury, FNflox/flox mice have been crossed with both PostnMerCreMer and Tcf21MerCreMer mice. Using FNflox/flox/PostnMerCreMer, we found that inducible ablation of FN in CFs post-I/R provided similar, albeit slightly lesser, myocardial protection post-I/R. Therefore, we hypothesize that inhibition of FN polymerization and/or ablation of CF-expressed FN attenuates cardiac remodeling by limiting pathologic CF activation and interstitial fibrosis (Figure 1). To test our hypothesis we propose the following aims: Aim 1: Determine the efficacy and specificity of inhibiting FN polymerization and/or ablating CF-restricted FN expression in attenuating the progression of HF. Aim 2: Determine the mechanism(s) of inhibiting FN polymerization and/or ablating FN expression in attenuating CF activation using primary mouse and human CFs. We believe this proposal, with FN inhibitory peptides validated in CF-restricted FN-/- mice as well as in human cardiac tissue, holds therapeutic promise for HF. !